Understanding inflammatory processes in ovine footrot to inform rational vaccine design

Lead Research Organisation: Moredun Research Institute
Department Name: Vaccines and Diagnostics

Abstract

Abstracts are not currently available in GtR for all funded research. This is normally because the abstract was not required at the time of proposal submission, but may be because it included sensitive information such as personal details.

Technical Summary

The aetiology of footrot is complex and multifactorial. Physical damage to the interdigital skin is required to initiate disease following exposure to damp conditions. The incidence of footrot could therefore increase in line with the climate change predictions of warmer and wetter summers in the UK. Bacterial replication in the damaged skin leads to interdigital dermatitis where the superficial epidermal layers are inflamed, damaged and slough off. This can progress to footrot with separation of the hoof horn from the underlying tissue. The essential bacterium causing footrot is Dichelobacter nodosus. However, the role of a second bacterium, Fusobacterium necrophorum, in the pathogenesis of footrot is unclear. The host response to the invasion of bacteria is characterised by inflammation of the epidermis and pressure in the hoof horn capsule. Consequently the hoof horn separates from the underlying tissue. The host immune response is a double-edged sword, while it is required for successful defence, if unbalanced it can lead to pathology. Given that separation of the hoof horn is characterised by immune-mediated inflammation, investigating how this is initiated and controlled will dramatically improve our understanding of footrot pathogenesis. Analysing the global host immune response to footrot-associated bacteria is a first step in understanding local inflammation. Together with an insight into the molecular epidemiology of D. nodosus and analysis of the correlation of sequence type with disease presentation, this will greatly enhance our understanding of the aetiology of footrot.
This proposal will help to inform the design and delivery of novel therapeutic and vaccine approaches, with the ultimate aim of facilitating strategic effective vaccine development. This proposal aligns with solutions to food security, reducing antibiotic usage in meat production and the potential for resistance, and providing a sustainable, secure supply of good quality food.

Planned Impact

Footrot is a bacterial infection of the interdigital skin of the sheep foot resulting in lameness, and is the greatest welfare and economic concern for sheep farmers and veterinarians worldwide. In England, more than 95% of sheep flocks have footrot, with a mean daily prevalence of ~10% and an estimated cost to the UK sheep industry of £24-84 million per annum. The results from this project will help to improve our understanding and management of footrot, a disease that impacts this industry and those concerned with animal welfare. It is therefore of relevance to the sheep industry (both pedigree and commercial sheep farmers), their consultants including veterinarians and policy makers, Farm Animal Welfare Committee and the RSPCA. We will ensure that our work has high visibility through peer-reviewed publications, trade publications and through presentations at scientific, veterinary and farmer meetings. We will continue work with EBLEX, the levy body for beef and sheep farmers, and transfer knowledge as appropriate throughout the project allowing interaction and promotion of results to key stakeholders. The results are also of potentially high media interest and could lead to a radical change in the approach to managing the foot health of sheep. The farming press (Farming Today, Farmer's Weekly) are routes to communicating the outputs of this project to the farming industry. In addition, outcomes will inform teaching practice and have the potential to change prevention and treatment of footrot.
This is a collaborative, multidisciplinary project and we envisage that the proposed research will benefit a wide range of academics working in diverse fields. It will benefit immunologists studying innate immunity to bacterial infections by elucidating host recognition and signalling responses to bacteria in a farm animal species of veterinary importance. It will also provide valuable tools (ovine TLR expression constructs, ovine skin fibroblast cell lines) to study other host - microbe interactions in sheep. This research will be of interest to scientists studying mechanisms of inflammation and immunopathology in response to infections. The generated and freely available mammalian transcriptome and pathogen genome and community profiling data will benefit the research community for diverse future research. In addition, the MLST analysis is web based portable system that will allow others to 'plug and play' with their own data, thus developing an ever growing database of such information on D. nodosus globally, enabling bespoke global adaptation of project findings.
The public have a strong interest in the health and welfare of animals that are farmed for human consumption. This includes concerns over the use of antibiotics to control disease in food animals and the impact of disease on productivity that ultimately influences the quality and purchase cost of meat. Footrot in sheep impacts on all of these areas and is therefore pertinent to the global Food Security agenda. The applicants have expertise in communicating their science to broad audiences beyond the academic arena. Drs Tötemeyer and Coffey are involved in outreach activities including Science Week, Sutton Trust Summer Schools and a STEM ambassador aiming to communicate veterinary science to pupils, including pupils from widening participation backgrounds. Dr Tötemeyer also reviewed for the Wikivet project. Prof Entrican and Dr Wattegedera have communicated their science to farmers at The Royal Highland Show and open days at Moredun Research Institute. They have presented to the public at The Edinburgh International Science Festival and have delivered immunology workshops to school teachers addressing the learning objectives relating to immunology and infectious disease within the new Curriculum for Excellence.

Publications

10 25 50
 
Description 1. Development of a bacterial typing method. Bacterial species exist in many different subtypes, which can be seen by variations in their genes. These differences are thought to help the bacteria survive and spread which presents an additional challenge for vaccine development. Considering this, we have developed a new method for identifying subtypes of the bacterium that principally causes footrot in sheep, Dichelobacter nodosus. This new typing scheme identifies variations in several selected genes, allowing for subtle differences to be detected. The application of this method on bacteria isolated from within the U.K. has shown that each farm seems to have a specific collection of bacterial subtypes which differs from others, even those in close geographic proximity. Also, whilst in other countries where populations comprise of one or two subtypes the U.K. is highly variable making design of an efficacious difficult. Blanchard et al 2018, DOI 10.3389/fmicb.2018. 00551.

2. Understanding the bacterial population during the progression of ovine footrot. Although D. nodosus is the bacterium that principally causes footrot, there are other factors and bacteria that are associated with disease. Progression from the initial publication: A distinct bacterial dysbiosis associated skin inflammation in ovine footrot (Maboni et al 2017, DOI 10.1038/srep45220): We have identified bacterial populations in the tissue and on the skin surface and how those change in footrot. The use of Dual RNASeq has allowed us to identify the transcriptional profile for the host showing that genes relating to inflammation and chronic wounds are significantly higher expresses in footrot affected feet, while genes relating to the barrier function of the skin are significantly lower expressed compared to healthy feet from the same sheep.
3. In vitro infection models to investigate host response to footrot. We have developed a novel 3D skin explant model to study anaerobic bacterial infection such as footrot (Maboni et al 2017, DOI: 10.3389/fcimb.2017.00404). We have isolated primary ovine fibroblasts and keratinocytes from tissue samples to determine cellular host response. The host response of those primary cells to D. nodosus includes the expression of proinflammatory cytokines and metalloproteases. Transfection studies suggest a role for TLRs in the recognition of D. nodosus.
Exploitation Route Our new typing scheme and the associated database will be of use to researchers interested in understanding genetic exchange between footrot bacteria and how the bacteria evolve. The information will also be valuable to researchers interested in finding new vaccines to prevent footrot. The influence of microbiome communities on disease development and progression is very topical. The new information we are generating on the microbiome in footrot will be of interest to researchers who are aiming to draw associations between 'good' and 'bad' bacteria in the feet of healthy and diseased sheep. This could ultimately be linked to studies on the use of antibiotics in treatment of disease and the effect on the microbiome.
Sectors Agriculture

Food and Drink

 
Title A tiered Multi-Locus Sequence Typing (MLST) scheme for the Footrot causing pathogen Dichelobacter nodosus 
Description MLST uses DNA sequence variation in a set of genes required for basic cellular maintenance, to define allelic differences, and has become the gold standard for population analyses of bacterial pathogens. The main benefits of MLST are that it is portable and highly reproducible. The analysis is also automated through server based databases removing the bias potentially incorporated through different users and the complexity of data analysis. Sequence types are created by the generation of an allelic profile. This is a series of numbers based on novel sequence variation present in the seven alleles, for example the first loci combination for isolate one will be designated the starting profile (1-1-1-1-1-1-1). Any subsequent variation within an allele will generate allele two (e.g. 1-1-2-1-1-1-1) and so on for all unique loci for each isolate. With numerous Dichelobacter nodosus (D. nodosus) isolates publically available in the NCBI sequence read archive (SRA ID: ERP005873) the development of a tiered MLST scheme was undertaken with the aim of developing a robust typing method to define STs, which would be accessible to all, regardless of budget or experience. 
Type Of Material Improvements to research infrastructure 
Year Produced 2017 
Provided To Others? Yes  
Impact This study has addressed the lack of knowledge on the global relatedness of D. nodosus and interestingly highlights the lack of recombination at many levels of analysis. The will allow future researchers to add their isolates to the database to further inform the population structure of D. nodosus potentially leading to advances in determining vaccine targets. 
URL https://pubmlst.org/dnodosus/
 
Description ARC Dissemination Event 2016 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact Attendance at ARC dissemination event attended by scientists and industry where the project was presented and discussed.
Year(s) Of Engagement Activity 2016
 
Description Attendance at ARC Dissemination Event 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact Attendance at BBSRC ARC Dissmenination Event, Birmingham, 30-31 October 2019, where the project was presented by the lead PI (Sabine Totemeyer, University of Nottingham). There were discussions about future potential collaborative projects during the networking sessions at the event.
Year(s) Of Engagement Activity 2017
 
Description Attendance by PI at Animal Health Research Club Third Dissemination Event 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Presentations of projects funded within the BBSRC Animal Health Research Club (Academic Researchers and Industry). Discussion of data and results and projected plans for future work.
Year(s) Of Engagement Activity 2016
 
Description Final ARC Project Meeting 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Study participants or study members
Results and Impact A final meeting of the 'Understanding Inflammatory Processes in Ovine Footrot to Inform on Rational Vaccine Design' project team (BB/M012085/1 & BB/M011941/1) was held at the University of Nottingham on 22 February 2019 in association with the related ARC Project 'Is Multistrain Infection by Dichelobacter nodosus Important in the Severity of Footrot and Management of Disease?' (BB/M012980/1). The aims of the meeting were to review the outcomes of the research and identify potential future projects. The meeting was attended by a total of 20 investigators, researchers and students who had contributed to the projects. There were 12 presentations, followed by a round-table discussion which focussed on 'knowns' and 'unknowns' of ovine footrot. This generated a number of ideas which were captured and will be used to formulate future funding applications.
Year(s) Of Engagement Activity 2019
 
Description Final ARC Project Meeting 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Study participants or study members
Results and Impact A final meeting of the 'Understanding Inflammatory Processes in Ovine Footrot to Inform on Rational Vaccine Design' project team (BB/M012085/1 & BB/M011941/1) was held at the University of Nottingham on 22 February 2019 in association with the related ARC Project 'Is Multistrain Infection by Dichelobacter nodosus Important in the Severity of Footrot and Management of Disease?' (BB/M012980/1). The aims of the meeting were to review the outcomes of the research and identify potential future projects. The meeting was attended by a total of 20 investigators, researchers and students who had contributed to the projects. There were 12 presentations, followed by a round-table discussion which focussed on 'knowns' and 'unknowns' of ovine footrot. This generated a number of ideas which were captured and will be used to formulate future funding applications.
Year(s) Of Engagement Activity 2019
 
Description Final BBSRC ARC Dissemination Event 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact Attended the final BBSRC ARC Dissemination Event 11-12th December in Manchester, UK. The project was presented to ARC partners and there was discussion about the findings. Plans were put in place for a final project meeting in Nottingham in early 2019 that would be attended by members of the other ARC Footrot Project (University of Warwick).
Year(s) Of Engagement Activity 2019
 
Description Public Education Event at Edinburgh International Science Festival 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Edinburgh International Science Festival public event. Practical demonstration and hands-on activity for schoolchildren to learn immunology techniques and the importance of vaccination to prevent infectious diseases.
Year(s) Of Engagement Activity 2017
 
Description Veterinary Immune Toolbox Meeting, Pirbright 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact A workshop was held at The Pirbright Institute to discuss national and international efforts to develop the Veterinary Immune Toolbox, which relates directly to the objectives of this project. A presentation was delivered by Gary Entrican on the history of global Veterinary Immune Reagent Development initiatives and a report was written which can be found at the hyperlink below.
Year(s) Of Engagement Activity 2017
URL http://www.vetvaccnet.ac.uk/publications/veterinary-immunology-toolbox-meeting-uk-veterinary-vaccino...
 
Description Veterinary Immune Toolbox Workshop 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact A Workshop on the Veterinary Immune Toolbox was held at the Veterinary Vaccinology Annual Conference in Stirling on 17 January 2018. The objective was to identify gaps in capability and make plans to develop web-based resources on immune reagent availability and function. A report on the conference can be found at the hyperlink below.
Year(s) Of Engagement Activity 2018
URL http://www.vetvaccnet.ac.uk/news/2018/01/uk-veterinary-vaccinology-network-conference-2018-report